Dengue (DEN) is the most important arthropod-borne viral infection of humans with about 100 million cases and 25,000 deaths annually, threatening over 3.5 billion people worldwide. The dengue viruses cause dengue fever (DF), dengue hemorrhagic fever (DHF), and dengue shock syndrome (DSS), and are endemic throughout the world's subtropical and tropical regions. To date, there is no effective vaccine to prevent against DF and no drug treatment for the disease. Dengue infection is caused by one of four different RNA viruses: dengue type 1 (DEN-1), DEN-2, DEN-3 and DEN-4. For a dengue vaccine to be safe and effective, it must be capable of neutralizing all four of the dengue viruses. Inviragen's tetravalent DEN vaccine (DENVax) consists of the live attenuated DEN-2 PDK-53 virus and three chimeras expressing the structural genes (prM and E) from DEN-1, DEN-3 and DEN-4, and retaining the genetic alterations responsible for the safety of the original DEN-2 vaccine. We have demonstrated the safety and efficacy of DENVax in AG129 mice and monkeys. In these studies we have identified tetravalent formulations that induce neutralizing antibodies to all four DENV serotypes. However, the responses to DENVax-4 were limited by interference from the other, more robust chimeras. In this proposal, we will test the hypothesis that the immune responses to the DENVax-4 vaccine construct can be further optimized through genetic manipulation of the DENVax-4 infectious clone. We propose to design and characterize a new chimeric DEN2/4 vaccine, and test the safety and efficacy of this vaccine in mice and monkeys. This proposal uniquely utilizes resources, facilities and reagents available at Inviragen Inc, CDC, and the University of Wisconsin-Madison. The development of an effective dengue vaccine represents an important approach to the prevention and control of this global emerging disease. Inviragen's long term goal is to develop a safe and effective dengue vaccine. PUBLIC HEALTH RELEVANCE: Dengue virus (DEN), a mosquito-borne RNA virus, is the most important arthropod-borne viral infection of humans with about 100 million cases and 25,000 deaths annually. We are developing a safe and effective tetravalent dengue vaccine that will protect against all four dengue serotypes. Such a vaccine would protect U.S. travelers from dengue infection, and significantly improve global public health as dengue threatens over 3.5 billion people worldwide.